1. Field of the Invention
The present invention relates to an efficient and inexpensive method of rapidly hydrolyzing oleaginous materials of all types into their constituent fatty acids and glycerol. This method will increase the production of fatty acids while reducing costs. The present invention also relates to an inherently immobilized lipase catalyst created from seeds which can be used to hydrolyze oleaginous materials with reduced reaction times despite use of modest temperatures.
2. Description of Related Art and Information
Fatty acids are important industrial chemicals used in the production of alkyd resins, dimer acids and dicarboxylic acids. The fatty acid industry has continuously attempted to increase free fatty acid production by searching for new methods of fat and oil hydrolysis. At the present time, in the fatty acid industry, hydrolysis of fats and oils is accomplished by a high-temperature steam treatment method known as the Colgate-Emery Steam Hydrolysis Process Brady, C., L. Metcalfek, D. Slaboszewski, and D. Frank, JAOCS, 65:917-921 (1988)!. This process operates with super-heated steam at 250.degree. C. and 50 atm. A two hour reaction results in a 97% yield of fatty acids. This high-temperature process is energy intensive, and causes extensive degradation of the produced fatty acids. Purification of the fatty acid product by means such as distillation is required prior to its recovery. In order to avoid degradation problems, some sectors of industry create fatty acids from vegetable oils by splitting such oils with a base followed by acidulation. This procedure does not, however, achieve complete splitting.
The use of enzymes to split fats is well known in the prior art. In 1948, A. W. Ralston reported that in 1890, Green and Sigmund, working independently, established the presence of a fat-splitting enzyme in castor beans, Ralston, A. W., "Fatty Acids and Their Derivatives," John Wiley & Sons, Inc., p. 275 (1948)!. These enzymes, known as lipases, function at an oil-water interface to hydrolyze fats and oils to fatty acids and glycerol. Many investigators have studied the enzymatic reaction and have encountered difficulty isolating the fatty acids from the heavy emulsion which is formed, Sonntag, N. O. V., Fat Splitting and Glycerol Recovery in Fatty Acids in Industry, edited by R. W. Johnson and E. Fritz, Marcel Dekker, Inc., New York, N.Y. (1989)!.
Hydrolysis of olive oil using Candida rugosa lipase has been reported, Linfield, W. M., D. J. O'Brien, S. Serota and R. A. Barauskas, J. Am. Oil Chem. Soc., 61:1067-1071, (1984)!. However, successful hydrolysis required prior treatment of the oil with bleaching earth.
Lipase-catalyzed hydrolysis of soybean oil has been reported. However, nearly complete lipolysis required the use of two lipases, as each lipase alone resulted in only partial splitting, Park, Y. K., G. M. Pastore and M. M. deAlmeida, J. Am. Oil Chem. Soc., 65:252-254, (1988)!.
It has been reported that some enzymes retain their activity in organic solvents Zaks, A., and A. M. Klibanov, Proceedings of the National Academy of Sciences, 82:3192-3196 (1985)!. Non-immobilized lipases from Candida rugosa, Rhizomucor miehei and porcine pancreas were shown to be catalytically active in organic solvents containing a trace of water for aminolysis, oximolysis and various esterification reactions. However, these enzyme systems were neither shown nor taught to be useful for hydrolytic reactions.
The use of lipases in non-aqueous solvents has been reviewed independently, by Gillis, A., JAOCS, 65:846-852 (1988); Klibanov, A. M., TIBS, 14:141-144 (1989); and Wong, C. H., Science, 244:1145-1152 (1989). In the majority of applications, these enzymes have been used to catalyze reactions such as the synthesis of ester bonds which are thermodynamically unfavorable in an aqueous medium. The lipase catalyzed formation of fatty amides from fatty acid methyl esters has also been reported, Bistline, R. G., Jr., A. Bilyk, and S. H. Feairheller, JAOCS, 68:95-98 (1991)!.
It has also been shown that lipases are lipolytically active in organic solvents. Use of fungal lipases in organic solvents, Bilyk et al., JAOCS, 68:320-323, (1991)!, achieved hydrolysis of a variety of fats and oils at moderate temperatures and in a relatively short time. Yield of fatty acids was limited to 76%, but with the addition of substantial amounts of substituted amines, yields of approximately 95% were obtained.
Lee and Hammond, in their article "Oat (Avena sativa) Caryopses as a Natural Lipase Bioreactor," J. Am. Oil. Chem. Soc., 67:761-765, (1990), reported the initial rate of hydrolysis of coconut oil and castor oil relative to that of soybean oil, by whole, dehulled oat seeds. The reaction system consisted of caryopses wetted with water and immersed in a hexane and oil mixture. Sometimes the mixtures were gently stirred. Although it was possible to achieve up to 90% lipolysis, this required 58 days and three batches of oat seeds. Only a 10% conversion was achieved after 4 days. They postulated that the slow rate of hydrolysis may be due to inhibition of the enzyme by glycerol, a product of the reaction. This reference does not disclose a rapid method of oil or animal fat splitting. Nor does the reference disclose a method of recycling or regeneration of the enzymes.
In order to conserve energy and obtain light colored fatty acids, some companies have investigated industrial enzymatic fat splitting using processes which involve the mixing of fats with lipase, agitating for 2 to 4 days, and subsequently isolating the fatty acids and glycerol, Meito Sangyo Col, Ltd., Jap. Pat. 79, 95, 607, Chem. Abstracts, 91, 21299069, (1979)!. For example, Miyoshi Fat and Oil, using a lipase supplied by Meito Sangyo Co., splits fats at 32.degree. C., developed the capacity to produce 1000 metric tons of fatty acids per month. Meito Sangyo does not suggest or teach a rapid method of fat and oil splitting using a lipase catalyst found in seeds.
U.S. Pat. No. 5,032,515, "Hydrolysis Process of Fat or Oil," to Tanigake et al., is a method which describes the hydrolysis of fat or oil by the continuous or semi-continuous supplying of water and lipase while simultaneously withdrawing a solution containing the fatty acids and glycerol to maintain the glycerol concentration in a range of 10 to 40% by weight. Tanigake et al. use lipase from Candida cylindracea.
U.S. Pat. No. 4,865,978, Lipolytic Splitting of Fats and Oils, to Serota, uses a spiral heating/cooling coil and a special mixer having impeller blades and baffles to prevent mass swirling to enzymatically hydrolyze triglycerides.
A reduction in lipolytic reaction time and temperature would result in a substantial reduction in the fat inventories presently required for the production process and a concurrent decrease in energy consumption and cost. Since lipases are currently too expensive to only be used in a single batch process, the ability to reuse enzymes would enhance the attractiveness of enzymatic hydrolysis in an industrial setting.